Outdoor Large scale Microalgae consortium culture for biofuel production in South Africa: Overcoming adverse environmental effects on microalgal growth

In nature, microalgal blooms occur regularly and in contrast with laboratory cultivation procedures, these blooms are not axenic and this seems to add to the longevity and intensity of the bloom. For this reason a consortium of microalgae and bacteria is used at InnoVenton for large scale biomass cultivation. The biomass produced, has successfully been used to produce biocrude. Laboratory cultivation procedures also require a large energy input in terms of artificial lighting, heating and aeration making it a costly endeavour. However, this stringent control of the culture to avoid contamination and ensure optimal growth conditions is essential when cultivating the microalgae for medical and pharmaceutical applications. When culturing for a chemical application such as biofuel production, this is not the case, therefore allowing for the economical outdoor consortium cultivation approach employed at InnoVenton. In this study we investigated whether morning heating of the media will overcome low consortium growth rates experienced in winter and whether the use of glucose, ethanol and acetate will overcome biomass loss exhibited at night. The results showed that heating increased the growth rate relative to those measured in summer and that heating all day in winter did not induce better growth rates in comparison to only heating for an hour at sunrise. We show that all three carbon sources are efficient at overcoming biomass loss at night with glucose being the most effective. In conclusion, employing these two techniques, the same growth rate theoretically can be achieved year round with large scale outdoor cultivation. Keywords: consortium, microalgae, biocrud

Uptake, Growth, and Pigment Changes in Lemna minor L. Exposed to Environmental Concentrations of Cylindrospermopsin

Cylindrospermopsin (CYN)-producing cyanobacterial blooms such as Raphidiopsis, Aphanizomenon, Anabaena, Umezakia, and Lyngbya spp. are occurring more commonly and frequently worldwide. CYN is an environmentally stable extracellular toxin, which inhibits protein synthesis, and, therefore, can potentially affect a wide variety of aquatic biota. Submerged and floating macrophytes, as primary producers in oligotrophic habitats, are at risk of exposure and information on the effects of CYN exposure at environmentally relevant concentrations is limited. In the present study, we investigated CYN uptake in the floating macrophyte Lemna minor with exposure to reported environmental concentrations. The effects were evaluated in terms of bioaccumulation, relative plant growth, and number of fronds per day. Variations in the concentrations and ratios of the chlorophylls as stress markers and carotenoids as markers of oxidative stress defense were measured. With exposure to 25 μg/L, L. minor could remove 43% of CYN within 24 h but CYN was not bioaccumulated. Generally, the pigment concentrations were elevated with exposure to 0.025, 0.25, and 2.5 μg/L CYN after 24 h, but normalized quickly thereafter. Changes in relative plant growth were observed with exposure to 0.25 and 2.5 μg/L CYN. Adverse effects were seen with these environmentally realistic concentrations within 24 h; however, L. minor successfully recovered within the next 48–96 h

Translocation of the cyanobacterial toxin microcystin-LR into guttation drops of Triticum aestivum and remaining toxicity

Uptake of the commonly occurring cyanobacterial toxin microcystin-LR (MC-LR) into crop plants via spray irrigation has been demonstrated. As other hazardous compounds such as pesticides were shown to be transported within plants, it was essential to understand the transport and fate of MC-LR in plants and the risks posed to grazers and other consumers. Of specific interest was to investigate if MC-LR could be detected in guttation drops and the toxicity thereof. Triticum aestivum (wheat) seedlings were exposed to 100 μg L−1 MC-LR in two separate experiments during which guttation drops were collected at various time points. The plants of one experiment were sectioned to investigate MC-LR distribution to the various plant appendages via liquid chromatography-tandem mass spectrometry analysis. After exposure, MC-LR could be detected in the roots, stems, leaves, and the guttation drops. However, the guttation drops were not toxic to Daphnia. As the environmentally relevant toxin concentration used was not sufficient to promote mortality in Daphnia, the physiological effect in insects, which rely on guttation drops as a water source, remains unknown. Combined with other contaminants that insects may be exposed to, the additional MC-LR exposure could contribute to the overall toxicity through the “tears of death”.Peer reviewe

Photocatalytic degradation of microcystin-LR by modified high-energy {001} titanium dioxide: Kinetics and mechanism study of HF8

Background: Uniquely synthesised titanium dioxide (TiO2) with high-energy {001} exposed facets denoted HF8 was used for the photocatalytic degradation of microcystin-LR (MC-LR) under ultraviolet irradiation at 365 nm. Methods: The influence of various conditions including environmental pH, nutrient anions, TiO2 dose, and MC-LR concentration was studied, and concentration of MC-LR measured using liquid chromatography-tandem mass spectrometry. Results: Within 120 min, 72.6% of an environmentally relevant MC-LR concentration (120 µg/L) was degraded under pH conditions ranging from 3 to 11. Stability tests revealed no loss of TiO2 activity after four applications of the same dose, indicating its stability, reusability, and potential to be re-used for sustainable remediation of MC-LR in eutrophic waters. Mechanism studies suggested that the reaction obeyed the pseudo-first-order equation and that hydroxyl radicals are the major reactive intermediate contributing to the reaction. The structure elucidation of intermediates suggested that hydroxylation and bond cleavage between the Adda chain and Mdha site could be the initiation of reactions in the degradation of MC-LR by HF8 TiO2. Conclusion: The results present a fast, sustainable, and practical method using modified TiO2 to improve MC-LR remediation.Peer reviewe

Interspecies interactions between Microcystis aeruginosa PCC 7806 and Desmodesmus subspicatus SAG 86.81 in a co-cultivation system at various growth phases

In lakes, cyanobacterial blooms are frequently associated with green algae and dominate the phytoplankton community in successive waves. In the present study, the interactions between Microcystis aeruginosa PCC 7806 and Desmodesmus subspicatus were studied to clarify the probable ecological significance of algal secondary metabolites; focusing on the role of cyanotoxin ‘microcystin-LR’(MC-LR). A dialysis co-cultivation technique was applied where M. aeruginosa was grown inside and D. subspicatus was cultured outside of the dialysis tubing. The concentration of the intra- and extracellular MC-LR and the growth of two species were measured at different time points over a period of one month. Additionally, the growth of the two species in the culture filtrate of one another and the effect of the purified MC-LR on the growth of the green alga were studied. The results indicated that the co-existing species could affect each other depending on the growth phases. Despite the early dominance of D. subspicatus during the logarithmic phase,M. aeruginosa suppressed the growth of the green alga at the stationary phase, which coincided with increased MC production and release. However, the inhibitory effects of Microcystis might be related to its other extracellular metabolites rather than, or possibly in addition to, MCPeer reviewe

Desmodesmus subspicatus co-cultured with microcystin producing (PCC 7806) and the non-producing (PCC 7005) strains of Microcystis aeruginosa.

Although microcystins (MCs) are the most commonly studied cyanotoxins, their significance to the producing organisms remains unclear. MCs are known as endotoxins, but they can be found in the surrounding environment due to cell lysis, designated as extracellular MCs. In the present study, the interactions between MC producing and the non-producing strains of Microcystis aeruginosa, PCC 7806 and PCC 7005, respectively, and a green alga, Desmodesmus subspicatus, were studied to better understand the probable ecological importance of MCs at the collapse phase of cyanobacterial blooms. We applied a dialysis co-cultivation system where M. aeruginosa was grown inside dialysis tubing for one month. Then, D. subspicatus was added to the culture system on the outside of the membrane. Consequently, the growth of D. subspicatus and MC contents were measured over a 14-day co-exposure period. The results showed that Microcystis negatively affected the green alga as the growth of D. subspicatus was significantly inhibited in co-cultivation with both the MC-producing and -deficient strains. However, the inhibitory effect of the MC-producing strain was greater and observed earlier compared to the MC-deficient strain. Thus, MCs might be considered as an assistant factor that, in combination with other secondary metabolites of Microcystis, reinforce the ability to outcompete co-existing species.Peer reviewe

Still challenging : the ecological function of the cyanobacterial toxin microcystin – What we know so far

Microcystins (MCs) are the most commonly studied cyanotoxins. While these past studies have mainly focused on the toxicity of MCs, the evolutionary history of life has shown that toxicity can be considered as an assigned role to MCs. Nowadays, there is a growing interest in understanding the importance of cyanotoxins in any of the physiological processes or beyond at the ecological level. This review evaluates variously proposed intracellular and extracellular functions of MCs and how they benefit the producing cyanobacterium. However, the strain-specific and divergent laboratory and field results obtained to date have made it difficult to generalize.Peer reviewe

Assessment of microplastic pollution : occurrence and characterisation in Vesijarvi lake and Pikku Vesijarvi pond, Finland

In the last few years, several studies have investigated microplastics (MPs) in marine ecosystems, but data monitoring and assessing the occurrence in freshwater environments are still scarce. The present study aims to investigate the occurrence, distribution, and chemical composition of MP pollution in Vesijärvi lake and Pikku Vesijärvi pond close to the city of Lahti (Finland) in winter. Sediment, snow, and ice core samples were collected near the shore of these two aquatic systems. MPs were analysed and identified by a non-destructive method using Fourier transform infrared spectroscopy (FTIR) 2D imaging. The mean concentrations of MPs detected in sediment, snow, and ice samples were 395.5 ± 90.7 MPs/kg, 117.1 ± 18.4 MPs/L, and 7.8 ± 1.2 MPs/L, respectively. FTIR results showed the predominant abundance of microplastics, such as polyamides (up to 53.3%), polyethylene and polypropylene (up to 17.1%), and natural fragments such as cellulose (up to 45.8%) and wool (up 18.8%) in the same size range. The potential release of MPs arising from stormwaters and sport and recreational activities was evidenced.Peer reviewe

Protein association of β-N-methylamino-L-alanine in Triticum aestivum via irrigation

Bioaccumulation of several cyanotoxins has been observed in numerous food webs. More recently, the neurotoxic, non-proteinogenic amino acid -N-methylamino-L-alanine (BMAA) was shown to biomagnify in marine food webs. It was thus necessary to assess whether a human exposure risk via a terrestrial food source could exist. As shown for other cyanotoxins, spray irrigation of crop plants with cyanobacterial bloom-contaminated surface water poses the risk of toxin transfer into edible plant parts. Therefore, in the present study, we evaluated a possible transfer of BMAA via spray irrigation into the seeds of one of the world's most widely cultivated crop plants, Triticum aestivum. Wheat plants were irrigated with water containing 10 mu g L-1 BMAA until they reached maturity and seed-bearing stage (205days). Several morphological characteristics, such as germination rate, number of roots per seedling, length of primary root and cotyledon, and diameter of the stems were evaluated to assess the effects of chronic exposure. After 205days, BMAA bioaccumulation was quantified in roots, shoots, and mature seeds of T. aestivum. No adverse morphology effects were observed and no free intracellular BMAA was detected in any of the exposed plants. However, in mature seeds, protein-associated BMAA was detected at 217 +/- 150ng g FW-1; significantly more than in roots and shoots. This result demonstrates the unexpected bioaccumulation of a hydrophilic compound and highlights the demand to specify in addition to limit values for drinking water, tolerable daily intake rates for the cyanobacterial-neurotoxin BMAA.Peer reviewe

Fate of Enrofloxacin in Lake Sediment : Biodegradation, Transformation Product Identification, and Ecotoxicological Implications

Various pharmaceutical drugs are being detected in different environmental compartments such as surface waters, groundwater, and sediment; a major concern since they are biologically active substances which can interfere with biological systems affecting the native biota. Among these drugs, antimicrobials are especially worrisome mainly due to the development of bacterial resistance. The aims of this study were to investigate if enrofloxacin, an emergent antibiotic pollutant, could be biodegraded in lake sediment, identify its break down products and to determine if these products have antimicrobial properties or are toxic. Three biodegradation products were identified and the antibiotic susceptibility assay proved that the products formed did not display antibiotic effects. Ecotoxicity testing with green algae suggested that the degradation products do not cause adverse effects statistically. However, it is suggested that further investigations are needed to identify the mechanism of degradation and the microbes involved.Peer reviewe